Our goal is the development of cell separation methods for the specific isolation of immune cells, particularly for varieties of antigen-reactive cells (ARC) involved in cellular immune reactions, and for their subcellular fractionation in order to study the mechanisms involved in the development of immune reactivities and immune macromolecules. Populations of cells containing ARC from normal mice and from mice immunized with allogeneic tumor cells and normal cells are tested for binding to the cell surface antigens to target cells attached to insoluble supports. Nonadherent cells are tested for depletion of cytotoxic effector cells (CTL) by measurement of 51Cr release from target cells. Graft-versus-host (GVH) activity is measured by the Simonsen assay in neonatal mice. The mixed lymphocyte culture is used to test for stimulation of ARC by alloantigen in vitro, and for the generation of CTL from their precursors (CTLp). T cell subpopulations from thymus and spleen are prepared and characterized with specific reagents such as peanut agglutinin and antibodies to the LyT and CTL differentiation antigens. Surface molecules of target cells are isolated to test their binding to ARC.